Bacillus subtilis CcdA-defective mutants are blocked in a late step of cytochrome c biogenesis

Cytochromes of the c type contain covalently bound heme. In bacteria, they are located on the outside of the cytoplasmic membrane. Cytochrome c synthesis involves export of heme and apocytochrome across the cytoplasmic membrane followed by ligation of heme to the polypeptide. Using radioactive protoheme IX produced in Escherichia coli, we show that Bacillus subtilis can use heme from the growth medium for cytochrome c synthesis. The B. subtilis ccdA gene encodes a 26-kDa integral membrane protein which is required for cytochrome c synthesis (T. Schiött et al., J. Bacteriol. 179:1962-1973, 1997). In this work, we analyzed the stage at which cytochrome c synthesis is blocked in a ccdA deletion mutant. The following steps were found to be normal in the mutant: (i) transcription and translation of cytochrome c structural genes, (ii) translocation of apocytochrome across the cytoplasmic membrane, and (iii) heme transport from the cytoplasm to cytochrome polypeptide on the outer side of the cytoplasmic membrane. It is concluded that CcdA is required for a late step in the cytochrome c synthesis pathway.